Ethan Garner Seminar 03/14/13

lab running for ~6 months now.


  • bacterial cell organization (in rod like cells): ends, middle, equally spaced
  • ParM finds the ends by pushing — polymerizing until it starts bending.

How bacteria grow

  • cell wall holds shape. Digest this, get spherical bacteria.
  • proteins acting at a local scale cause organization at 5 um scale
  • MreB – structural similarity to Actin, polymerizes
  • Ftz – structurally similar to Tubulin, polymerizes (core sequence not recognizable)

Audience Discussion

  • chromosome organization / shape looks different in Bacillus than E coli.
  • might be imaging condition difference.

Back to Bacteria growth

  • helical structure seen. Believed to be helical cytoskeleton
  • Motion still happens in absence of polymer dynamics — not treadmilling
  • maybe cell wall synthesis drives the motion
  • antibiotics and genetics that prevent cell wall synthesis prevent this motion.
  • all components move at a similar velocity.
  • All components are oriented perpendicular to the long axis of the cell.
  • clockwise/non-clockwise motion is uncorrelated at all length scales.
  • right before everything freezes (under drug / removal of cell wall) get random motion.

How do disconnected units organize in a coherent motion?

  • Tag0, have cell walls but still round. components move in all directions.
  • can induce rod formation in by activating the native gene — quickly start forming rods.

what is the function of MreB?

  • Can polyermerize and depolymerize reversibly with a drug
  • Class A PBPs (involved in inserting new cell wall), either diffusive or stationary, does not show coherent motion like the rest of the complex.

How is this growth regulated?

  • more enzymatic sites or faster synthesis?
  • Rate of motion doesn’t change. Must add more components
  • New fluorescent amino-acids. incorporate into cell wall, label newly synthesized cell wall.
  • Functional fusion proteins substantial challenge: MreB fusion protein makes nice helices. — These don’t compliment and these don’t move.
  • In ecoli, fusions do compliment and move, but different constructs behave differently.
  • some fusion proteins polymerize and never leave the polymer. Natural MreB makes small
  • long linkers 30+ and non-dimerization domains behave more natively.

Final observation

  • lipid involved in flipping glycoprotein chains from inside to outside, when inhibited, leads to filament polymerization.
  • amount of precursor determines amount of mREB determines amount of cell wall synthesis.

Final discussion

  • not clear if it is sugar polymerization driving the motion or something else.

Q -> can you target the incorporation of the labeled amino-acids into particular proteins / what’s the chance of doing this with clever genetics in the future? A -> currently not sure how they incorporate in the first place. Concievable to make labeled proteins with the technique (but probably a ways off).

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